神经生长因子对小鼠葡聚糖硫酸钠结肠炎的作用及机制

发布时间：2018-03-23 07:00

  本文选题：神经生长因子　切入点：溃疡性结肠炎　出处：《兰州大学》2014年硕士论文　论文类型：学位论文

【摘要】：目的使用神经生长因子(Nerve growth factor,NGF)对葡聚糖硫酸钠(Dextran sulfate sodium,DSS)诱导的急性溃疡性结肠炎小鼠进行药物干预,探讨神经生长因子(NGF)对小鼠溃疡性结肠炎的作用及可能机制。 方法16只6-8周龄(18-20g)的雌性SPF级Balb/c小鼠自由饮用5%DSS蒸馏水溶液7天后继续饮用蒸馏水7天,ELISA法测定肠组织NGF含量随时间变化。将40只Balb/c小鼠,按照随机原则分为A,B,C,D四组,每组10只,设：A组,正常对照组； B组,模型组,又叫DSS组； C组,DSS+Ad-NGF组,简称Ad-NGF组; D组,DSS+Ad-0组,简称Ad-0组。除A组饮用蒸馏水外,其余三组小鼠自由饮用5%DSS蒸馏水溶液,进食标准饲料。第7天,四组小鼠禁食12小时,不限饮水,第8天,A,B组动物经肛灌注0.9%生理盐水100μ L-次,C组经肛注入100μ L含1.O×109CFU/mL Ad-NGF, D组注入100u L1.0x109CFU/mL, B, C, D组饮用的5%DSS蒸馏水溶液改为正常饮水,一直持续到第14天。每天定时记录各组小鼠的一般状态,大便性状,隐血／肉眼血便,行疾病活动度(DAI)评分,第14天脱颈处死小鼠,采集结肠组织,测量结肠长度,部分组织石蜡包埋,行HE染色和免疫组化染色,部分结肠标本放置于-80℃冰箱保存,比色法及ELISA法测定MPO及IL-10,IL-6,TNF-a,免疫印迹法测定CGRP变化。 结果结肠组织NGF的浓度从第1天到第5天逐渐增加(0.095±0.032pg/g,0.32+0.24pg/g结肠组织),第6天达到最高值(0.52±0.45pg/g结肠组织),然后从第7天开始逐渐从0.09±0.04pg/g结肠组织下降到对照水平；而在第10天,浓度(0.06±0.023pg/g)较对照组低。DAI和组织学评分在DSS组显著升高(P0.01),但在Ad-NGF直肠内给药后明显下降(P0.05),而AD-0组该数据无明显变化。与Ad-0组(9.15±0.21cm)相比, NGF能够减弱DSS致(10.33+0.36cm)结肠长度的缩短(对照组vsDSS组：12.67±0.38cmvs7.98±0.15cm,p0.01)。同样,NGF显著抑制了DSS诱导的炎性因子IL-6(DSS组vs Ad-NGF:304.57±3.19vs56.67±5.89, p0.05), TNF-a (115.89±6.91,69.14＝4.48, p0.05)的升高,但促进了IL-10的分泌(65.34±6.44,160.73±6.42,p0.05),Ad-0无此作用。NGF显著抑制了DSS诱导的结肠炎模型中的MPO浓度的增高,Ad-NGF组浓度低于DSS组和Ad-0组(p0.05),DSS组和Ad-0组浓度无明显差异。DSS显著抑制了肠组织CGRP的产生,而；补充NGF显著升高了CGRP的含量,Ad-0无此作用。 结论神经生长因子可减轻DSS诱导的小鼠结肠炎性损伤；抑制MPO和炎性细胞因子,促进IL-10和CGRP的分泌可能是其抗炎的机制。神经生长因子可能成为临床治疗溃疡性结肠炎一个新的选择。
[Abstract]:Objective to investigate the effect of nerve growth factor (NGF) on ulcerative colitis in mice induced by Dextran sulfate sodiumn (DSS) and its possible mechanism. Methods Sixteen female SPF grade Balb/c mice (6-8 weeks old) were randomly divided into four groups: 5%DSS distilled water solution for 7 days and then distilled water for 7 days. 40 Balb/c mice were randomly divided into four groups. Ten rats in each group were divided into two groups: group A, normal control group; group B, model group, also called DSS group; group C, DSS Ad-NGF group, referred to as Ad-NGF group; group D, DSS Ad-0 group, referred to as Ad-0 group. Except for group A, the other three groups of mice drank 5%DSS distilled water solution freely. On the 7th day, the mice in the four groups fasted for 12 hours, without restriction on drinking water. On the 8th day, 100 渭 L of 0.9% normal saline was injected into the anus of rats in group B and 100 渭 L of 109CFU/mL Ad-NGF was injected into the anus of group C, and the 5%DSS distilled water solution of group D was injected with 100u L 1.0 x 109 CFU / mL, B, C, D instead of normal drinking water. It lasted until the 14th day. The general state, defecation character, occult blood / naked blood stool, disease activity and DAI score were recorded regularly every day. The mice were killed by removing neck on the 14th day, the colonic tissue was collected and the colonic length was measured. Some tissues were embedded in paraffin wax and stained with HE and immunohistochemistry. Some colon specimens were stored in refrigerator at -80 鈩，

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